We propose to study tumor cell invasion in novel in vitro systems utilizing endothelial cell monolayers grown on top of connective tissue collagen gels or bovine lens capsule basement membrane. Homotypic (tumor cells only) and heterotypic (tumor cells and immunocytes from tumor bearing hosts) tumor spheroids will be studied. The metastatic variant mouse melanoma cell lines B16F1, B16F10, the rat metastatic variant hepatocarcinoma cell lines RT-7-4b, RT-7-4bL and the mouse mammary tumor variants for anchorage requirement for growth, +SA, -SA as well as a limited number of human mammary tumor cells (to be supplied by Dr. C. McGrath, Michigan Cancer Center, Detroit) will be used. Findings from the in vitro studies will be correlated with behavior of the rodent tumor cell lines in vivo, or with clinical and pathological histories in the case of human material. We will assay for tissue degradative enzymes of putative importance to invasion and extravasation. Purified substrates as well as basement membrane matrices will be used as substrates. The ability of the spheroids to form stable attachments to endothelial cell monolayers, to induce endothelial cell retraction and to degrade and invade basement membrane and connective tissue substrates will be assessed. The ability of a series of proteolytic enzyme inhibitors to interfere with these activities, in particular with endothelial cell retraction, will be studied also. We will devote particular attention to any enzymatic or other contribution of tumor associated macrophages and other immunocytes which acids in tumor cell invasion or extravasation from heterotypic spheroids.